(519f) Virus Filtration: Effect of Protein Fouling and Transmembrane Pressure

Virus filtration is extensively used in the downstream purification process for the production of protein-based biotherapeutics. Commercial virus filters can ensure robust viral clearance by removing more than 99.9% of viruses while providing nearly complete protein transmission. However the size of most therapeutic products, e.g., monoclonal antibodies, are only slightly smaller than the 20 nm parvoviruses, leading to a significant amount of protein fouling. The overall objective of this study was to obtain more fundamental insights into the fouling behavior and its effect on virus capture by direct visualization of fouled membranes using both confocal and electron microscopy.

Filtration experiments were conducted using a single-layer, highly asymmetric Viresolve® Pro membrane with the skin-side facing away from the feed (as per the manufacturer’s recommendation). Filters were challenged with fluorescently-labeled IgG and imaged by confocal laser scanning microscopy to determine the location of retained protein within the depth of the filter. Parallel experiments were performed by filtering a suspension of gold nanoparticles of different sizes through protein-fouled filters, with the membrane cross-section imaged using scanning electron microscopy to identify the location of individual captured nanoparticles.

IgG filtration leads to a significant decline in flux, with the extent of fouling being greatest at low transmembrane pressure. Confocal images of protein-fouled membranes show the presence of fluorescently-labeled IgG in a narrow band located immediately upstream of the filter exit, with greater fluorescent intensity observed at low transmembrane pressure (consistent with the increased flux decline). SEM images obtained after filtration of gold nanoparticles show a band of 20 nm particles located approximately 1 µm upstream of the filter exit. Protein fouling caused the location of the captured 20 nm gold particles to shift further away from the filter exit, likely due to constriction of the small pores in the Viresolve® Pro membrane. These results provide important insights into the factors controlling the performance of virus filters and the mechanisms governing protein fouling.